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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Expression of glycosylated human prolactin in HEK293 cells and related N-glycan composition analysis

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Author(s):
Silva, Felipe D. [1] ; Oliveira, Joao E. [1] ; Freire, Renan P. [1] ; Suzuki, Miriam F. [1] ; Soares, Carlos R. [1] ; Bartolini, Paolo [1]
Total Authors: 6
Affiliation:
[1] IPEN, CNEN SP, Biotechnol Ctr, Ave Prof Lineu Prestes 2242, BR-05508000 Sao Paulo - Brazil
Total Affiliations: 1
Document type: Journal article
Source: AMB EXPRESS; v. 9, n. 1 AUG 29 2019.
Web of Science Citations: 0
Abstract

Prolactin (PRL) is a hormone produced by the pituitary gland with innumerable functions, such as lactation, reproduction, osmotic and immune regulation. The present work describes the synthesis of hPRL in human embryonic kidney (HEK293) cells, transiently transfected with the pcDNA-3.4-TOPO (R) vector carrying the hPRL cDNA. A concentration of 20 mg/L, including glycosylated (G-hPRL) and non-glycosylated (NG-hPRL) human prolactin, was obtained, with 19% of G-hPRL, which is higher than that observed in CHO-derived hPRL ( 10%) and falling within the wide range of 5-30% reported for pituitary-derived hPRL. N-Glycoprofiling analysis of G-hPRL provided: (i) identification of each N-glycan structure and relative intensity; (ii) average N-glycan mass; (iii) molecular mass of the whole glycoprotein and relative carbohydrate mass fraction; (iv) mass fraction of each monosaccharide. The data obtained were compared to pituitary- and CHO-derived G-hPRL. The whole MM of HEK-derived G-hPRL, determined via MALDI-TOF-MS, was 25,123 Da, which is 0.88% higher than pit- and 0.61% higher than CHO-derived G-hPRL. The main difference with the latter was due to sialylation, which was sevenfold lower, but slightly higher than that observed in native G-hPRL. The ``in vitro{''} bioactivity of HEK-G-hPRL was fourfold lower than that of native G-hPRL, with which it had in common also the number of N-glycan structures. (AU)

FAPESP's process: 15/26058-0 - Glycoprofiling and N-glycosylation site occupancy of different human thyrotropin (hTSH) preparations of pituitary or recombinant origin
Grantee:Carlos Roberto Jorge Soares
Support type: Regular Research Grants
FAPESP's process: 17/50332-0 - Scientific, technological and infrastructure qualification in radiopharmaceuticals, radiation and entrepreneurship for health purposes
Grantee:Marcelo Linardi
Support type: Research Grants - State Research Institutes Modernization Program